During the past four years, we have shown that an aminosterol, MSI-1436, decreases body weight in rodents by inhibiting food intake and increasing thermogenesis. MSI-1436 is effective in diet-induced obese as well as leptin-deficient mice, and rapidly decreases plasma glucose and stimulates fatty acid oxidation in these models. Since MSI-1436 is more potent when injected in the cerebral ventricle and into the hypothalamic paraventricular nucleus, it is likely that the pharmacologic effects of this compound occur directly in the brain. Indeed, we have shown increased binding of MSI-1436 within the arcuate nucleus, and suppression of AGRP and NPY mRNA expression in mice, implicating neural pathways expressing these neuropeptides as MSI-1436 targets. Independent of weight loss, MSI-1436 alters the expression of enzymes involved in lipid and glucose metabolism, such as hypothalamic AMP kinase (AMPK) and hepatic fatty acid synthase (FAS). Furthermore, agouti (Ay/a) mice are less sensitive than wild-type animals to MSI-1436 and the response to MSI-1436 is inhibited by SHU9119, consistent with the hypothesis that the melanocortin system is a critical pathway in the central mode of action of MSI-1436. We have recently identified a mouse hypothalamic cell line (GT1-7) in which MSI-1436, at submicromolar concentrations, enhanced the agonist activity of a-MSH, suggesting a possible mechanism by which MSI-1436 might influence the melanocortin pathway. This proposal will further explore the hypothesis that MSI-1436 acts via central mechanisms. Aim 1 will evaluate the effects of pharmacologic blockade or genetic deletion of melanocortin 4 receptor (MC4R) on the ability of MSI-1436 to inhibit feeding and stimulate thermogenesis via the hypothalamic paraventricular nucleus. We will explore whether corticotropin-releasing hormone (CRH) acts downstream of MC4R to mediate the effects of MSI-1436 on feeding and thermogenesis. Aim 2 will attempt to resolve the mechanism by which MSI-1436 enhances the in vitro binding and signaling via MC4R in GT1-7 hypothalamic neurons. Finally, Aim 3 will determine whether the rapid actions on glucose and lipid metabolism are mediated through MC4R and AMPK in the hypothalamus. Understanding of the central neuronal actions MSI-1436 will provide novel insights on the anti-obesity and insulin sensitizing actions of this aminosterol.